We have successfully transmitted the malignant phenotype from tumor to morphologically normal cells via transfection with discrete fragments of tumor DNA. This experimental approach has allowed us to detect the presence of transforming genes in a variety of human tumors and human tumor cell lines. They include carcinomas of the bladder, colon, gall bladder, liver, lung and pancreas, as well as in a fibrosarcoma and an embryonal rhabdomyosarcoma. The transforming gene present in T24 cells in a human bladder carcinoma cell line has been isolated by molecular cloning techniques. We have demonstrated that it represents the human homologue of the oncogene present in the Harvey and BALB strains of murine sarcoma viruses. Finally, we have established that the T24 bladder oncogene is identical to its normal allele present in all human cells except for a single nucleotide, the basic chemical unit of the DNA molecule. Thus, a single mutational event is responsible for the malignant activation of this human transforming gene.